The systematic wrapping of an axon by insulating myelin sheaths is a remarkable event in the development of the vertebrate central nervous system. Despite the importance of myelin for the rapid conduction of action potentials, little is known about its molecular mechanism. We have been using the rat optic nerve as a simple model system to study myelination. When purified rat retinal ganglion cells are cultured together with purified oligodendrocyte precursor cells (OPCs) little myelination occurs. However, when the pharmacological inhibitor of gamma-secretase, DAPT, is added to the culture medium, robust myelination occurs within 3 days, providing us with a simple culture system to study myelination. In this application, we will focus on the hypothesis that CNS myelination is normally controlled by a gamma-secretase substrate. First, we will test whether DAPT promotes myelination in culture by inhibiting gamma-secretase activity in neurons or in oligodendrocytes and investigate whether gamma-secretase activity within the developing optic nerve normally helps to control myelination. Second, we will test 3 specific candidate signaling pathways, known to be regulated by gamma-secretase, that have previously been implicated as potential controllers of myelination: the Jagged1-Notch1 pathway, Neuregulin-erbB4 pathway, and N-cadherin-mediated adhesion. Third, we will determine whether DAPT promotes myelination by enhancing local axon-glial interactions or by inducing oligodendrocytes to differentiate to a myelinating stage. We will perform time-lapse microscopy to ask fundamental questions about how oligodendrocytes myelinate axons. Finally, we will study the gene changes that accompany DAPT-induced myelination. Together, we hope that these experiments will provide a better understanding of the molecular mechanisms that control CNS myelination. Understanding how myelination is regulated may suggest new ways of enhancing remyelination after injury or diseases such as optic neuritis and Multiple Sclerosis.